Given that late stage melanoma has been notoriously difficult to treat it is urgent to harness the immune system to resist metastases. In murine and human models, tumor regression can be mediated by tumor-specific cytolytic T cells via adoptive immunotherapy or by directly inducing CTLs in vivo. Our laboratory has demonstrated that an effective way to generate antigen-specific CTLs is by using dendritic cells [DCs], potent antigen presenting cells in the blood, as "natural" adjuvants. We have perfected a method for obtaining these cells in large numbers and in potent immunostimulatory form from blood monocytes. In vivo, monocyte-derived antigen-pulsed DCs dramatically boost CTL responses in humans, and eradicated or markedly decrease tumor load in murine models. Moreover, DCs prepared from patients with breast or ovarian cancer antigens via cross-presentation. In preclinical studies, we propose to study stage III melanoma patients and exploit the immunostimulatory capacity of DCs to detect helper and CTL responses in vitro to tumor associated antigens expressed by melanoma cell lines, and to identify novel antigenic epitopes. Advantage will be taken of [a] new assays that measure responses at the single cell level e.g. ELISPOT, tetramer and intracellular cytokine assays and [b] recall assays where DCs are used as APCs. Based on the information gained we will first immunize stage III individuals with antigen-pulsed autologous DCs. Immune T cells from patients, along with T cell clones to defined melanoma cells will be compared with peptide-pulsed DCs in their capacity to immunize stage IV patients in a randomized bi-center trial. We hypothesize that vaccination with DCs bearing melanoma antigens, especially dying cells, will lead to elevations in CTLp and helper cells, rapid mobilization of effector CTLs and lasting immunity.